Binary combination of epsilon-poly-L-lysine and isoeugenol affect progression of spoilage microbiota in fresh turkey meat, and delay onset of spoilage in Pseudomonas putida challenged meat.

Proliferation of microbial population on fresh poultry meat over time elicits spoilage when reaching unacceptable levels, during which process slime production, microorganism colony formation, negative organoleptic impact and meat structure change are observed. Spoilage organisms in raw meat, especially Gram-negative bacteria can be difficult to combat due to their cell wall composition. In this study, the natural antimicrobial agents ε-poly-L-lysine (ε-PL) and isoeugenol were tested individually and in combinations for their activities against a selection of Gram-negative strains in vitro. All combinations resulted in additive interactions between ε-PL and isoeugenol towards the bacteria tested. The killing efficiency of different ratios of the two antimicrobial agents was further evaluated in vitro against Pseudomonas putida. Subsequently, the most efficient ratio was applied to a raw turkey meat model system which was incubated for 96 h at spoilage temperature. Half of the samples were challenged with P. putida, and the bacterial load and microbial community composition was followed over time. CFU counts revealed that the antimicrobial blend was able to lower the amount of viable Pseudomonas spp. by one log compared to untreated samples of challenged turkey meat, while the single compounds had no effect on the population. However, the compounds had no effect on Pseudomonas spp. CFU in unchallenged meat. Next-generation sequencing offered culture-independent insight into population diversity and changes in microbial composition of the meat during spoilage and in response to antimicrobial treatment. Spoilage of unchallenged turkey meat resulted in decreasing species diversity over time, regardless of whether the samples received antimicrobial treatment. The microbiota composition of untreated unchallenged meat progressed from a Pseudomonas spp. to a Pseudomonas spp., Photobacterium spp., and Brochothrix thermosphacta dominated food matrix on the expense of low abundance species. We observed a similar shift among the dominant species in meat treated with ε-PL or the antimicrobial blend, but the samples differed markedly in the composition of less abundant species. In contrast, the overall species diversity was constant during incubation of turkey meat challenged with P. putida although the microbiota composition did change over time. Untreated or ε-PL treated samples progressed from a Pseudomonas spp. to a Pseudomonas spp. and Enterobacteriaceae dominated food matrix, while treatment with the antimicrobial blend resulted in increased relative abundance of Hafnia spp., Enterococcaceae, and Photobacterium spp. We conclude that the blend delayed the onset of spoilage of challenged meat, and that all antimicrobial treatments of unchallenged or challenged meat affect the progression of the microbial community composition. Our study confirms that the antimicrobial effects observed in vitro can be extrapolated to a food matrix such as turkey meat. However, it also underlines the consequence of species-to-species variation in susceptibility to antimicrobials, namely that the microbial community change while the CFU remains the same. Addition of antimicrobials may thus prevent the growth of some microorganisms, allowing others to proliferate in their place.

Hafnia psychrotolerans sp. nov., isolated from lake water.

A psychrotolerant, Gram-stain-negative, motile, aerobic, peritrichous bacterium, strain DJC1-1(T), was isolated from Lake Dajiaco, Tibetan Plateau, China. The strain was negative for citrate utilization, lipase activity and α-glucosidase, but positive for the Voges-Proskauer reaction and N-acetyl-ß-glucosaminidase. 16S rRNA gene sequence analysis indicated that Hafnia paralvei ATCC 29927(T), Hafnia alvei ATCC 13337(T), Serratia grimesii DSM 30063(T) and Serratia plymuthica DSM 4540(T) were the closest relatives of strain DJC1-1(T), with similarities of 97.76, 96.80, 97.71 and 97.58 %, respectively. The DNA G+C content of strain DJC1-1(T) was 53.9 mol%. The predominant fatty acids were C16 : 0 and C17 : 0 cyclo. Based on these characteristics, strain DJC1-1(T) can be assigned to the genus Hafnia. In DNA-DNA hybridization tests, strain DJC1-1(T) shared 50.6, 35.1, 36.5 and 18.1 % DNA-DNA relatedness with the type strains of H. paralvei, H. alvei, S. grimesii and S. plymuthica, respectively. The growth temperature ranged from 0 to 40 °C, with optimum growth at 15 °C. Physiological and biochemical tests differentiated strain DJC1-1(T) from the type strains of recognized species of the genus Hafnia. Therefore, strain DJC1-1(T) is identified as representing a novel species of the genus Hafnia, for which the name Hafnia psychrotolerans sp. nov. is proposed. The type strain is DJC1-1(T) ( = JCM 30077(T) = CGMCC1.12806(T)).

Conversion of glycerol to 1,3-propanediol by Citrobacter freundii and Hafnia alvei - newly isolated strains from the Enterobacteriaceae.

In this study, nearly 4000 bacterial strains from the family of Enterobacteriaceae isolated from different environments were screened for ability to convert glycerol to 1,3-propanediol (1,3-PD). The aim of the research was to isolate 1,3-PD producers from the natural environment, identify and characterize the best isolates. Three selective media were tested to usefulness in the isolation of bacteria from the family Enterobacteriaceae. Only, 28% of examined isolates could synthesize 1,3-PD from glycerol. 1,3-PD producing bacteria were identified by API 20E tests and 16S rRNA sequences to be Klebsiella pneumoniae, Klebsiella oxytoca, Citrobacter freundii and Hafnia alvei. It is the first time, when the fermentation glycerol to 1,3-PD by H. alvei was investigated. The selected strains (C. freundii AD119 and H. alvei AD27) were analyzed on a bioreactor scale under constant pH value 7.0 at temperature of 30°C and 37°C. After 40h in batch fermentation, H. alvei AD27 produced 11.3g/L of 1,3-PD at 37°C. For C. freundii AD119, the best results were obtained at temperature of 30°C. After 24h of fermentation, the 1,3-PD concentration reached above 23 g/L of 1,3-PD.